Starch synthase IIIa and starch branching enzyme IIb-deficient mutant rice line ameliorates pancreatic insulin secretion in rats: screening and evaluating mutant rice lines with antidiabetic functionalities.

Diabetes mellitus is a metabolic disease spreading worldwide that has been reported to worsen the development and progression of other diseases (cancer, vascular diseases and dementia). To establish functional rice lines with anti-postprandial hyperglycaemic effects, we developed mutant rice lines, which lack one or two gene(s) related to starch synthesis, and evaluated their effects. Powder of mutant rice lines or other grains was loaded to rats fasted overnight (oral grain powder loading test). Incremental area under time-concentration curves (iAUC) were calculated with monitored blood glucose levels. Rice lines with anti-postprandial hyperglycaemic effects were separated by cluster analysis with calculated iAUC. A double mutant rice #4019 (starch synthase IIIa (ss3a)/branching enzyme IIb (be2b)), one of the screened mutant rice lines, was fed to Goto-Kakizaki (GK) rats, an animal model for type 2 diabetes, for 5 weeks. Plasma levels of C-peptide, a marker of pancreatic insulin secretion, were measured with ELISA. For in vitro study, a rat pancreatic cell line was cultured with a medium containing rat serum which was sampled from rats fed #4019 diet for 2 d. After 24-h of incubation, an insulin secretion test was performed. Through the oral rice powder loading test, seven rice lines were identified as antidiabetic rice lines. The intake of #4019 diet increased plasma C-peptide levels of GK rats. This result was also observed in vitro. In rat serum added to cell medium, ornithine was significantly increased by the intake of #4019. In conclusion, the mutant rice #4019 promoted pancreatic insulin secretion via elevation of serum ornithine levels.

Development and evaluation of rice giant embryo mutants for high oil content originated from a high-yielding cultivar 'Mizuhochikara'.

Rice bran oil is a byproduct of the milling of rice (Oryza sativa L.). It offers various health benefits and has a beneficial fatty acid composition. To increase the amount of rice bran as a sink for triacylglycerol (TAG), we developed and characterized new breeding materials with giant embryos. To induce mutants, we treated fertilized egg cells of the high-yielding cultivar 'Mizuhochikara' with N-methyl-N-nitrosourea (MNU). By screening M2 seeds, we isolated four giant embryo mutant lines. Genetic analysis revealed that the causative loci in lines MGE12 and MGE13 were allelic to giant embryo (ge) on chromosome 7, and had base changes in the causal gene Os07g0603700. On the other hand, the causative loci in lines MGE8 and MGE14 were not allelic to ge, and both were newly mapped on chromosome 3. The TAG contents of all four mutant lines increased relative to their wild type, 'Mizuhochikara'. MGE13 was agronomically similar to 'Mizuhochikara' and would be useful for breeding for improved oil content.

Effect of organizational factors and work environments on newly graduated nurses' intention to leave.

AIM: The purpose of this study was to investigate intention to leave in relation to organizational factors and work environments among newly graduated nurses (NGN). METHODS: A questionnaire was delivered to NGN (n = 762) from baccalaureate programs as well as diploma schools from 19 R-hospitals (R-hospitals were designated as "public medical institutions" by the 1951 Medical Law) in 2010. Spearman's rank correlation coefficients and Mann-Whitney U-test were used to test statistical significance. RESULTS: Only 148 questionnaires were returned for a return rate of 19.4%. The authors found that 8.1% of respondents had a highly to extremely likely desire to leave nursing. Intention to leave was significantly associated with Nursing Work Index total (P < 0.01), social support (P < 0.001), work barriers (P < 0.001), commitment to workplace (P < 0.001), job satisfaction (P < 0.001), and burnout (P < 0.001). Those who graduated from R-schools run by the R-organization had significantly positive impressions of their organizations and significantly lower intentions to leave compared to other graduates. CONCLUSION: The study results suggest that NGN intention to leave the workplace and nursing predict actual early intention to leave. In order to reduce burnout and turnover, support from colleagues and supervisors appears to be essential in organizational and psychosocial work environments for NGN to experience job satisfaction and be committed to their workplace. The results of this study also suggest that a longitudinal study is needed to reveal the long-term effects of organizational factors and work environments on NGN and how they impact role transition and adaptation of the NGN to professional practice roles.

Mucosal immune responses in W/W(v) and Sl/Sl(d) mutant mice.

In order to identify potential unanticipated side reactions and immune responses, the evaluation of candidate vaccines should include immunization of the murine model of the disease in question and mutant animals, as well as normal laboratory animals. We employed WBB6F(1)-W/W(v) and WBB6F(1)-Sl/Sl(d) mutant mice, which are genetically mast cell deficient and lack intestinal pacemaker activity due to a severe deficiency in interstitial cells of Cajal. Antigen-specific mucosal and systemic immune responses in the mutant and congenic normal mice were induced by intranasal or intragastric immunization with ovalbumin (OVA) plus cholera toxin as an adjuvant. It was found that the levels of the OVA-specific humoral immune response in the mucosal and systemic tissues of the mutant mice immunized intranasally were roughly equivalent to those of the congenic normal mice. In contrast, the specific humoral immune response in the intragastrically immunized mutant mice was greater than that observed in the congenic normal mice. Unexpectedly, the titers of OVA-specific IgA antibodies and total IgA antibodies in the fecal extracts of both intranasally and intragastrically immunized mutant mice were significantly lower than in those of the congenic normal mice. Although the detailed mechanisms leading to these differences remain unclear, the unexpected immune responses observed in the gastrointestinal tracts of the mice in this study may be related to an abnormality of gastrointestinal motility. Our data therefore suggest that studies using mutant mice and physiological assessments should be carried out during mucosal vaccine development.

The indonesian variants of CRF33_01B: Near-full length sequence analysis.

Cocirculation of subtype B and CRF01_AE in Southeast Asia has led to the establishment of new recombinant forms. In our previous study, we found five samples suspected of being recombinants between subtype B and CRF01_AE, and here, we analyzed near full-length sequences of two samples and compared them to known CRFs_01B, subtype B, and CRF01_AE. Five overlapped segments were amplified with nested PCR from PBMC DNA, sequenced, and analyzed for genome mosaicism. The two Indonesian samples, 07IDJKT189 and 07IDJKT194, showed genome-mosaic patterns similar to CRF33_01B references from Malaysia, with one short segment in the 3' end of the p31 integrase-coding region, which was rather more similar to subtype B than CRF01_AE, consisting of unclassified sequences. These results suggest gene-specific continuous diversification and spread of the CRF33_01B genomes in Southeast Asia.

Demonstration of iodine K-edge imaging by use of an energy-discrimination X-ray computed tomography system with a cadmium telluride detector.

An energy-discrimination K-edge X-ray computed tomography (CT) system is useful for increasing the contrast resolution of a target region by utilizing contrast media. The CT system has a cadmium telluride (CdTe) detector, and a projection curve is obtained by linear scanning with use of the CdTe detector in conjunction with an X-stage. An object is rotated by a rotation step angle with use of a turntable between the linear scans. Thus, CT is carried out by repetition of the linear scanning and the rotation of an object. Penetrating X-ray photons from the object are detected by the CdTe detector, and event signals of X-ray photons are produced with use of charge-sensitive and shaping amplifiers. Both the photon energy and the energy width are selected by use of a multi-channel analyzer, and the number of photons is counted by a counter card. For performing energy discrimination, a low-dose-rate X-ray generator for photon counting was developed; the maximum tube voltage and the minimum tube current were 110 kV and 1.0 microA, respectively. In energy-discrimination CT, the tube voltage and the current were 60 kV and 20.0 microA, respectively, and the X-ray intensity was 0.735 microGy/s at 1.0 m from the source and with a tube voltage of 60 kV. Demonstration of enhanced iodine K-edge X-ray CT was carried out by selection of photons with energies just beyond the iodine K-edge energy of 33.2 keV.

Current HIV type 1 molecular epidemiology profile and identification of unique recombinant forms in Jakarta, Indonesia.

HIV infection is a major problem in Indonesia. The number of people living with HIV has been increasing from year to year, especially among injecting drug users (IDUs). Since there were only limited data about molecular epidemiology profiles of HIV/AIDS in Indonesia, a cross-sectional study involving 208 HIV-1-seropositive individuals was conducted in 2007 in Jakarta. The majority of participants were 16-30 years of age (64.9%) and 74.5% were male. The most frequent risk factor was injecting drug use (IDU) (45.7%) followed by heterosexual transmission (34.1%). Phylogenetic analysis of gag (p17 and p6) and env C2V3 regions showed 200 (96.2%) of 208 DNA samples were CRF01_AE and only 3 (1.4%) were subtype B. Five samples (2.4%) indicated discordant subtypes between the three aforementioned regions: three of them showed unique CRF01_AE/B recombination patterns in 2.3-kbp nucleotide sequences (from p17 to part of RT), including one sample showing similarity to CRF33_01B, reported previously in Malaysia. This study shows the current predominant subtype is CRF01_AE in every risk group, with a decreasing number of pure subtype B, and the first identification of CRF01_AE/B recombinant forms among HIV-1-seropositive Indonesians.

Co-administration of cholera toxin and apple polyphenol extract as a novel and safe mucosal adjuvant strategy.

Although native cholera toxin (CT) is an extremely effective adjuvant, its toxicity prevents its use in humans. We report here that apple polyphenol extract (APE), obtained from unripe apples, reduces CT-induced morphological changes and cAMP accumulation. Based upon this finding, we have attempted to design a novel, effective and safe mucosal vaccine by using CT with several dosages of APE as nasal adjuvants. Mice nasally immunized with OVA plus CT and an optimal dosage of APE showed significantly reduced levels of inflammatory responses as well as total and OVA-specific IgE antibodies when compared with mice given without APE. However, levels of both mucosal and systemic OVA-specific antibody responses were maintained. Further, APE significantly down-regulated accumulation of CT in the olfactory nerves and epithelium. In summary, an optimal dosage of APE would take full advantage of mucosal adjuvanticity of native CT without any toxicity for application in humans.

Neuronal apoptosis and inflammatory responses in the central nervous system of a rabbit treated with Shiga toxin-2.

BACKGROUND: Shiga toxins (Stxs) are the major agents responsible for hemorrhagic colitis and hemolytic-uremic syndrome (HUS) during infections caused by Stx-producing Escherichia coli (STEC) such as serotype O157:H7. Central nervous system (CNS) involvement is an important determinant of mortality in diarrhea associated-HUS. It has been suggested that vascular endothelial injuries caused by Stxs play a crucial role in the development of the disease. The current study investigates the relationship between the cytotoxic effects of Stxs and inflammatory responses in a rabbit brain treated with Stx2. METHODS: In a rabbit model treated with purified Stx2 or PBS(-), we examined the expression of the Stx receptor globotriaosylceramide (Gb3)/CD77 in the CNS and microglial activation using immunohistochemistry. The relationship between inflammatory responses and neuronal cell death was analyzed by the following methods: real time quantitative reverse transcriptase (RT)-polymerase chain reaction (PCR) to determine the expression levels of pro-inflammatory cytokines, and the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) method to detect apoptotic changes. RESULTS: Gb3/CD77 expression was detected in endothelial cells but not in neurons or glial cells. In the spinal cord gray matter, significant levels of Gb3/CD77 expression were observed. Severe endothelial injury and microvascular thrombosis resulted in extensive necrotic infarction, which led to acute neuronal damage. Conversely, in the brain, Stx receptor expression was much lower. The observed neuropathology was less severe. However, neuronal apoptosis was observed at the onset of neurological symptoms, and the number of apoptotic cells significantly increased in the brain at a later stage, several days after onset. Microglial activation was observed, and tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta mRNA in the CNS parenchyma was significantly up-regulated. There was significant overexpression of TNF-alpha transcripts in the brain. CONCLUSION: This study indicates that Stx2 may not directly damage neural cells, but rather inflammatory responses occur in the brain parenchyma in response to primary injury by Stx2 in vascular endothelial cells expressing Gb3/CD77. These findings suggest that neuroinflammation may play a critical role in neurodegenerative processes during STEC infection and that anti-inflammatory intervention may have therapeutic potential.

[Serum anti-lPS antibody production in an outbreak of enterohemorrhagic Escherichia coli O157 infection among schoolchildren].

Although there have been many reports of the usefulness of serodiagnosis of enterohemorrhagic Escherichia coli (EHEC) O157, the serotype of the bacteria detected and the increase in anti-LPS antibody have not always been consistent. In this study we investigated the diagnostic significance of measurements of anti-LPS antibody by ELISA in an outbreak of O157 infection among schoolchildren in whom the bacteriological test findings were clarified and the age groups were uniform. The anti-LPS antibody titer was measured in 31 patients (77 serum samples) in an outbreak of EHEC O157 : H7 infection (220 children infected) that occurred in a primary school in Morioka in 1996. The anti-O157 LPS antibody positivity rates of IgM, IgG, and IgA were 98.7%, 85.7%, and 98.7%, respectively. Between the time the meal that caused the outbreak and 19 days later, anti-O157 LPS IgM antibody and IgA antibody were detected in all patients. The specificity was investigated using control serum, and the specificity of IgM, IgG, and IgA was 93.5%, 93.5%, and 97.2%, respectively. Some samples contained antibodies against O111 and O26 LPS, but the titers were lower than the anti-O157 antibody titer. The anti-O111 antibody titer and anti-O26 antibody titer were highly correlated, suggesting that they were crossreactive antibodies for O157 LPS. No significant correlation was found between differences in clinical manifestations and the anti-O157 LPS antibody titer in this O157 outbreak in schoolchildren. It was clarified that an increase in anti-LPS antibody was found to support the diagnosis of mild cases of 0157 infection infection as well as severe cases.

A silkworm larvae plasma test for detecting peptidoglycan in cerebrospinal fluid is useful for the diagnosis of bacterial meningitis.

The silkworm larvae plasma (SLP) test has been established based on a cascade reaction triggered by either peptidoglycan or (1, 3)-beta-D-glucan to produce melanin. We applied this test to the diagnosis of bacterial meningitis. Cerebrospinal fluid (CSF) obtained from patients with bacterial meningitis due to gram-positive bacteria, gram-negative bacteria, or fungi, showed positive reactions to the test. In contrast, CSF from patients with viral meningitis or noninfectious illnesses gave negative reactions. Therefore, this test seems to be useful for diagnosis of bacterial and fungal meningitis. When this test was used together with two types of limulus tests, an endotoxin-specific test, and a conventional test, meningitis was further characterized as gram-positive, gram-negative or fungal meningitis. The SLP test requires a computerized instrument for quantitative colorimetric measurement. A qualitative alternative of this test also can be accomplished by visually observing the darkening color. Thus, this method can be applied for simple and rapid diagnosis of meningitis.

First report of CRF03_AB recombinant HIV type 1 in injecting drug users in Ukraine.

It has been reported that subtypes A throughout H of HIV-1 are circulating in the former Soviet Union. In this sequence note, we analyzed the genetic prevalence of HIV-1 among injecting drug users (IDUs) in Ukraine. The subjects studied included two individuals from Kiev and six individuals from Simferopol', the latter located in the Crimean Peninsula. We found that one of eight IDUs was infected with a CRF03 gagA/envB recombinant HIV-1 and was from Simferopol', whereas the others were infected with HIV-1 subtype A. There combinant was closely related to other A/B recombinants reported previously, and had silent mutations Inthe V3 region, the same as other envB strains of HIV-1 circulating among mDUs in the former Soviet Union. The data supported reports that the Russian AIB recombinant HIV-1 was probably from Ukraine. This is the first report of a CRF03 gagA/envB recombinant HIV-1 found in Ukraine.